scholarly journals Improved Cold-resistant Performance in Transgenic Grape (Vitis vinifera L.) Overexpressing Cold-inducible Transcription Factors AtDREB1b

HortScience ◽  
2009 ◽  
Vol 44 (1) ◽  
pp. 35-39 ◽  
Author(s):  
Wanmei Jin ◽  
Jing Dong ◽  
Yuanlei Hu ◽  
Zhongping Lin ◽  
Xuefeng Xu ◽  
...  
Keyword(s):  
Transgenic Plants ◽  
Vitis Vinifera ◽  
Blot Analysis ◽  
Freezing Point ◽  
Cold Treatment ◽  
Cauliflower Mosaic Virus ◽  
Vitis Vinifera L ◽  
35S Promoter ◽  
Transgenic Lines ◽  
H 26

Dehydration response element binding (DREB)1b is a cold-inducible transcription factor in Arabidopsis thaliana. DREB1b driven by cauliflower mosaic virus 35S promoter was genetically introduced into grape Vitis vinifera L. cv. Centennial Seedless through Agrobacterium-mediated transformation for improving its cold resistance and exploring new genetic breeding approaches to obtain cold-resistant cultivars. In this study, Southern blot analysis showed the DREB1b gene was integrated into the transgenic grapevines with one to two copies. Northern blot analysis showed the presence of DREB1b transcripts in the independent transgenic lines 3, 5, 6, and 7. Further characterization of transgenic grapevines confirmed that both electrolyte leakage conductivity and the freezing point of the transgenic plants were lower than those of wild-type plants. After the cold treatment at –4 °C for 12 h, 26% of transgenic plants wilted among which 95% plants recovered once being placed under the condition of temperature 22 to 25 °C. However, subjected to the same treatment, 98% of nontransgenic plants wilted and only 2% recovered. Our results lead to the conclusion that activity of DREB1b in the transgenic grape could significantly improve its resistance to cold stress.

scholarly journals Constitutive Expression of Pea Defense Gene DRR206 Confers Resistance to Blackleg (Leptosphaeria maculans) Disease in Transgenic Canola (Brassica napus)

1999 ◽  
Vol 12 (5) ◽  
pp. 410-418 ◽  
Author(s):  
Yaping Wang ◽  
Goska Nowak ◽  
David Culley ◽  
Lee A. Hadwiger ◽  
Brian Fristensky
Keyword(s):  
Brassica Napus ◽  
Transgenic Plants ◽  
Leptosphaeria Maculans ◽  
Constitutive Expression ◽  
Single Copy ◽  
Cauliflower Mosaic Virus ◽  
Adult Plant ◽  
35S Promoter ◽  
Transgenic Lines

To identify genes effective against the blackleg fungus Leptosphaeria maculans (Phoma lingam), we have transformed canola (Brassica napus) with four pea (Pisum sativum) genes under constitutive control by the cauliflower mosaic virus 35S promoter: PR10.1, chitinase, DRR206, and defensin. Transgenic lines containing single-copy T-DNA insertions for each gene were screened for both seedling (cotyledonary) and adult plant resistance. Lines for which pea DRR206 mRNA was expressed showed decreased disease scores, compared with non-expressing transgenic lines. Transgenic plants expressing pea defensin showed a slight enhancement of resistance, while for PR10 and chitinase transgenics there was little or no enhancement of resistance. Resistance to L. maculans cosegregated with DRR206 transgenes. Extracts from DRR206 and defensin transgenic plants inhibited fungal germination in vitro. DRR206 transgenic plants also demonstrated decreased hyphal growth at inoculation sites. While the precise function of DRR206 remains to be determined, these results suggest that it does play an important role in defense against fungi.

Aberrant promoter methylation occurred from multicopy transgene and SU(VAR)3 - 9 homolog 9 (SUVH9) gene in transgenic Nicotiana benthamiana

2012 ◽  
Vol 39 (9) ◽  
pp. 764 ◽  
Author(s):  
Gi-Ho Lee ◽  
Seong-Han Sohn ◽  
Eun-Young Park ◽  
Young-Doo Park
Keyword(s):  
Gene Silencing ◽  
Nicotiana Benthamiana ◽  
Fluorescent Protein ◽  
Cauliflower Mosaic Virus ◽  
Camv 35S Promoter ◽  
35S Promoter ◽  
Camv 35S ◽  
Histone Deacetylase 1 ◽  
Transgenic Lines ◽  
Green Fluorescent

The chemical modification of DNA by methylation is a heritable trait and can be subsequently reversed without altering the original DNA sequence. Methylation can reduce or silence gene expression and is a component of a host’s defence response to foreign nucleic acids. In our study, we employed a plant transformation strategy using Nicotiana benthamiana Domin to study the heritable stability of the introduced transgenes. Through the introduction of the cauliflower mosaic virus (CaMV) 35S promoter and the green fluorescent protein (GFP) reporter gene, we demonstrated that this introduced promoter often triggers a homology-dependent gene-silencing (HDGS) response. These spontaneous transgene-silencing phenomena are due to methylation of the CaMV 35S promoter CAAT box during transgenic plant growth. This process is catalysed by SU(VAR)3–9 homologue 9 (SUVH9), histone deacetylase 1 (HDA1) and domains rearranged methylase 2 (DRM2). In particular, we showed from our data that SUVH9 is the key regulator of methylation activity in epigenetically silenced GFP transgenic lines; therefore, our findings demonstrate that an introduced viral promoter and transgene can be subject to a homology-dependent gene-silencing mechanism that can downregulate its expression and negatively influence the heritable stability of the transgene.

scholarly journals Expression of thaumatin, a new type of alternative sweetener in rice

2020 ◽  
Vol 48 (3) ◽  
pp. 1276-1291
Author(s):  
Shahina AKTER ◽  
Md. Amdadul HUQ ◽  
Yu-Jin JUNG ◽  
Kwon-Kyoo KANG
Keyword(s):  
Transgenic Rice ◽  
Oryza Sativa L ◽  
Plasmid Vector ◽  
Single Copy ◽  
Cauliflower Mosaic Virus ◽  
Pcr Analysis ◽  
35S Promoter ◽  
Alternative Sweetener ◽  
Sweet Proteins ◽  
Transgenic Lines

  Sweet proteins are the natural alternative to the artificial sweeteners as well as flavor enhancers. Among other sweet protein, thaumatin protein was isolated from Thaumatococcus daniellii Benth plant fruit. In this study, pinII Ti plasmid vector was constructed with thaumatin gene, where thaumatin was placed under the control of the duel cauliflower mosaic virus 35S promoter into rice (Oryza sativa L. var. japonica cv. ‘Dongjinbyeo’) by Agrobacterium-mediated transformation to generate transgenic plants. Thirteen plant lines were regenerated and the transgenic rice lines were confirmed by different molecular analysis. The genomic PCR result revealed that all of the plant lines were transgenic. The single copy and intergenic plant lines were selected by Taqman PCR analysis and FST analysis, respectively. Expression of thaumatin gene in transgenic rice resulted in the accumulation of thaumatin protein in the leave. Thaumatin protein was also accumulated in leave of T1 generation. Sensory analysis result suggested that the thaumatin protein expressing transgenic lines exerted sweet tasting activity. These results demonstrated that thaumatin was expressed in transgenic rice plants.

scholarly journals Paxillus involutus Forms an Ectomycorrhizal Symbiosis and Enhances Survival of PtCOMT-modified Betula pendula in vitro

2008 ◽  
Vol 57 (1-6) ◽  
pp. 235-242 ◽  
Author(s):  
H. Tiimonen ◽  
T. Aronen ◽  
T. Laakso ◽  
P. Saranpää ◽  
V. Chiang ◽  
...  
Keyword(s):  
Populus Tremuloides ◽  
Betula Pendula ◽  
Root Formation ◽  
Lateral Roots ◽  
Cauliflower Mosaic Virus ◽  
35S Promoter ◽  
Paxillus Involutus ◽  
Transgenic Lines ◽  
G Ratio

Abstract The ability of the PtCOMT (caffeate/5-hydroxyferulate O-methyltransferase from Populus tremuloides L.) - modified Betula pendula Roth. lines to form symbiosis with an ectomycorrhizal (ECM) fungus Paxillus involutus Batsch Fr. was studied in vitro. Lignin precursor gene PtCOMT was introduced into two B. pendula clones under the control of the cauliflower mosaic virus 35S promoter or the promoter of the sunflower polyubiquitin gene UbB1. Of the four transgenic lines, one 35SPtCOMT line (23) had a decreased syringyl/guaiacyl (S/G) ratio of root lignin, and two UbB1-PtCOMT lines (110 and 130) retarded root growth compared to the control clone. Both control clones and all transgenic lines were able to form ECMs with P. involutus, but the transgenic lines differed from the controls in the characteristics of the ECMs. The number of lateral roots covered with fungal hyphae and/or development of a Hartig net (HN) were reduced in line 23 with a decreased S/G ratio, and in lines 110 and 130 with slower root formation and changed root morphology, respectively. However, line 23 benefited more from the inoculation in lateral root formation than the control, and in lines 110 and 130 the percentage of viable plants increased most due to inoculation. The results show that B. pendula plants genetically transformed with the lignin gene PtCOMT could form mycorrhizal symbiosis regardless of changes in either the root S/G ratio or development. The benefits of the symbiosis were variable even in the closed in vitro system, and dependent on the clone or transgenic line and the ECM fungal symbiont.

scholarly journals An amphibian antimicrobial peptide variant expressed in Nicotiana tabacum confers resistance to phytopathogens1

2003 ◽  
Vol 370 (1) ◽  
pp. 121-127 ◽  
Author(s):  
Donatella PONTI ◽  
M. LUISA MANGONI ◽  
Giuseppina MIGNOGNA ◽  
Maurizio SIMMACO ◽  
Donatella BARRA
Keyword(s):  
Nicotiana Tabacum ◽  
Transgenic Plants ◽  
Antimicrobial Peptide ◽  
Plant Pathogens ◽  
Rana Esculenta ◽  
Cauliflower Mosaic Virus ◽  
35S Promoter ◽  
Skin Secretions ◽  
Fungal Phytopathogens ◽  
Micro Organisms

Esculentin-1 is a 46-residue antimicrobial peptide present in skin secretions of Rana esculenta. It is effective against a wide variety of micro-organisms, including plant pathogens with negligible effects on eukaryotic cells. As a possible approach to enhance plant resistance, a DNA coding for esculentin-1, with the substitution Met-28Leu, was fused at the C-terminal end of the leader sequence of endopolygalacturonase-inhibiting protein, under the control of the cauliflower mosaic virus 35S promoter region, and introduced into Nicotiana tabacum. The antimicrobial peptide was isolated from the intercellular fluids of healthy leaves of transgenic plants, suggesting that it was properly processed, secreted outside cells and accumulated in the intercellular spaces. The morphology of transgenic plants was unaffected. Challenging these plants with bacterial or fungal phytopathogens demonstrated enhanced resistance up to the second generation. Moreover, transgenic plants displayed insecticidal properties.

Expression of a Chicken Ovalbumin Gene in Three Lucerne Cultivars

1991 ◽  
Vol 18 (5) ◽  
pp. 495 ◽  
Author(s):  
HE Schroeder ◽  
MRI Khan ◽  
WR Knibb ◽  
D Spencer ◽  
TJV Higgins
Keyword(s):  
Transgenic Plants ◽  
Alternative Interpretation ◽  
Cauliflower Mosaic Virus ◽  
Vector System ◽  
35S Promoter ◽  
Restriction Enzyme Digestion ◽  
Flanking Sequence ◽  
Gene Encoding ◽  
Transformed Plants ◽  
Chicken Ovalbumin

Routine procedures have been developed for the transformation of lucerne (Medicago sativa cv. Rangelander) with foreign genes using the Agrobacterium tumefaciens binary vector system and for the regeneration of transgenic plants from tissue culture, via somatic embryogenesis. Lucerne transformation was carried out with a gene encoding neomycin phosphotransferase (npt), which conferred resistance to the antibiotic kanamycin, together with a cDNA clone encoding chicken ovalbumin which was modified for expression in plant cells. The ovalbumin cDNA protein coding sequence was combined with the cauliflower mosaic virus 35S promoter and the nopaline synthase 3' flanking sequence to make a chimeric ovalbumin gene. A DNA construct containing both these genes was transferred to lucerne, and ovalbumin was detected in leaves of regenerated plants using protein immunoblots. Pulse-chase labelling experiments and analysis of leaves from the top to bottom of the transformed plants indicated that ovalbumin, once formed, was stable in the leaves of transgenic lucerne. A wide variation in ovalbumin level was frequently observed in plants regenerated from multiple embryos on a single transformed callus. This variation correlated with changes in the restriction enzyme digestion pattern of the ovalbumin DNA from the transgenic plants. These results indicate that each transformed callus may have arisen from more than one transformation event. An alternative interpretation is that the callus may have arisen from a single transformed cell but during cell proliferation the DNA in some cells may have undergone rearrangement prior to embryogenesis. Transformation and regeneration procedures were also developed for two Australian commercial cultivars of lucerne. Although the frequency of recovery of transformed plants was lower than with cv. Rangelander, these protocols open the way for a relatively rapid

scholarly journals A DNAβ Associated with Tomato Yellow Leaf Curl China Virus Is Required for Symptom Induction

2004 ◽  
Vol 78 (24) ◽  
pp. 13966-13974 ◽  
Author(s):  
Xiaofeng Cui ◽  
Xiaorong Tao ◽  
Yan Xie ◽  
Claude M. Fauquet ◽  
Xueping Zhou
Keyword(s):  
Transgenic Plants ◽  
Stop Codon ◽  
Blot Hybridization ◽  
Southern Blot Hybridization ◽  
Tomato Yellow Leaf ◽  
Cauliflower Mosaic Virus ◽  
Start Codon ◽  
35S Promoter ◽  
Yellow Leaf ◽  
Leaf Curl

ABSTRACT We report here that all 25 isolates of Tomato yellow leaf curl China virus (TYLCCNV) collected from tobacco, tomato, or Siegesbeckia orientalis plants in different regions of Yunnan Province, China, were associated with DNAβ molecules. To investigate the biological role of DNAβ, full-length infectious clones of viral DNA and DNAβ of TYLCCNV isolate Y10 (TYLCCNV-Y10) were agroinoculated into Nicotiana benthamiana, Nicotiana glutinosa, Nicotiana. tabacum Samsun (NN or nn), tomato, and petunia plants. We found that TYLCCNV-Y10 alone could systemically infect these plants, but no symptoms were induced. TYLCCNV-Y10 DNAβ was required, in addition to TYLCCNV-Y10, for induction of leaf curl disease in these hosts. Similar to TYLCCNV-Y10, DNAβ of TYLCCNV isolate Y64 was also found to be required for induction of typical leaf curl diseases in the hosts tested. When the βC1 gene of TYLCCNV-Y10 DNAβ was mutated, the mutants failed to induce leaf curl symptoms in N. benthamiana when coinoculated with TYLCCNV-Y10. However, Southern blot hybridization analyses showed that the mutated DNAβ molecules were replicated. When N. benthamiana and N. tabacum plants were transformed with a construct containing the βC1 gene under the control of the Cauliflower mosaic virus 35S promoter, many transgenic plants developed leaf curl symptoms similar to those caused by a virus, the severity of which paralleled the level of βC1 transcripts, while transgenic plants transformed with the βC1 gene containing a stop codon after the start codon remained symptomless. Thus, expression of a βC1 gene is adequate for induction of symptoms of viral infection in the absence of virus.

Transcriptional silencing of RNAi constructs against nematode genes in Arabidopsis

Nematology ◽  
2013 ◽  
Vol 15 (5) ◽  
pp. 519-528 ◽  
Author(s):  
Tina Kyndt ◽  
Hongli Ji ◽  
Bartel Vanholme ◽  
Godelieve Gheysen
Keyword(s):  
Transcriptional Silencing ◽  
Housekeeping Genes ◽  
Cauliflower Mosaic Virus ◽  
Heterodera Schachtii ◽  
Camv 35S Promoter ◽  
35S Promoter ◽  
Expression Levels ◽  
Camv 35S ◽  
General Decline ◽  
Transgenic Lines

In this research, Arabidopsis thaliana plants were transformed with hairpin constructs targeting cyst nematode (Heterodera schachtii) genes, driven by the cauliflower mosaic virus (CaMV) 35S promoter: two housekeeping genes (the splicing factor Hs-U2AF and the vacuolar Hs-H+ATPase) and one candidate effector gene (the ubiquitin extension protein Hs-ubi). Expression of the dsRNA appeared to be extremely variable between and within homozygous T3 lines and even between tissues. Infection experiments showed up to 50% reduction in nematode infection for some transgenic lines. The results varied not only between lines containing the same construct but also between independent repetitions of the experiment. Further focusing on the Hs-U2AF-RNAi lines revealed large variations and a general decline of construct expression levels over the generations. Bisulphite sequencing of a 197 bp part of the CaMV 35S promoter revealed substantial methylation in this region and a negative correlation between the methylation level and expression of the hairpin construct. Taken together, our results show that host-generated RNAi can suffer from high levels of transcriptional silencing of the construct, leading to varying expression levels within and between transgenic lines.

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